P989: NEWLY IDENTIFIED ROLES FOR PIEZO1 MECHANOSENSOR IN CONTROLLING NORMAL MEGAKARYOCYTE DEVELOPMENT AND IN PRIMARY MYELOFIBROSIS
Authors: Vittorio Abbonante, Iris A. Karkempetzaki, Christian A. DI Buduo, Nasi Huang, Daniele Cattaneo, Christina Ward, Shinobu Matsuura, Alessandra Iurlo, Katya Ravid, Alessandra Balduini
Journal: HemaSphere (2023)

Topic: 15. Myeloproliferative neoplasms - Biology & Translational Research Background: Primary Myelofibrosis (PMF) is a clonal Myeloproliferative Neoplasm (MPN) characterized by profound alterations of bone marrow architecture and mechanical features. Megakaryocytosis with atypical megakaryocytes (Mks) is a hallmark of PMF. Differently to Essential Thrombocythemia (ET), characterized by persistent thrombocytosis, PMF patients often develop thrombocytopenia with disease progression. The most frequent driver mutation in MPN is the JAK2V617F. Mechanical forces are important modulators of Mk development and function. We previously demonstrated that Mks express the Transient Receptor Potential Vanilloid 4 (TRPV4) cation channel, which activation, by the bone marrow soft environment, determines an increase of platelet formation. PIEZO1 is a newly characterized mechanosensitive cation channel expressed in different tissues. Its role in controlling the proliferation and differentiation of different cell types in response to mechanical stimuli has been proven, while its function in Mks is not known. Aims: We investigated a possible involvement of PIEZO1 in determining the typical Mk alterations found in PMF by studying its expression and function in samples derived from a JAK2V617F mouse model and in samples derived from patients affected by JAK2V617F MPNs. Methods: The role of PIEZO1 was investigated in mouse and human healthy samples by pharmacological treatments with the specific agonist YODA1 and the inhibitor GsMTx4. Samples derived from JAK2V617F mice and from patients affected by JAK2V617F PMF and ET were used to explore the role of PIEZO1 in megakaryopoiesis and proplatelet formation. Quantitative capillary-based electrophoresis was used to study PIEZO1 expression in primary Mks isolated from bone marrow aspirates of MPN patients. Results: We found that PIEZO1 is expressed during megakaryopoiesis and that its activation negatively impacts on Mk maturation as shown by the effect on cell size, ploidy, expression of the late marker CD42b, and ability of Mk to extend proplatelets. These features recapitulate the main defects observed in PMF Mks. Mks derived from mice bearing the JAK2V617F mutation showed increased PIEZO1 expression, compared to WT controls. Pharmacological activation of PIEZO1 significantly augmented the already elevated number of immature Mks, and sharply reduced the number of mature ones. Accordingly, Mks isolated from bone marrow aspirates of JAK2V617F PMF patients showed increased PIEZO1 expression compared to ET patients. Most importantly, PIEZO1 expression in bone marrow Mks inversely correlated with patient peripheral blood platelet count. The size, ploidy, expression of CD42b, and proplatelet formation of Mks, from thrombocytopenic JAK2V617F+ PMF patients, were rescued when differentiated in the presence of the PIEZO1 inhibitor GsMTx4. Another main feature of PMF Mks, a constitutively increased ERK1/2 activation, was normalized by GsMTx4 treatment. Summary/Conclusion: Together, our data show that PIEZO1 might serve as break to full Mk maturation and platelet formation under normal conditions, and its upregulation in PMF Mks further aggravates hallmarks of this disease. Keywords: Myeloproliferative disorder, Megakaryocyte differentiation, Bone marrow microenvironment, Proplatelet